Late appearance of resistance to tumor rechallenge following cryosurgery: A study in an experimental mammary tumor of the rat

Resistance to tumor challenge following surgical and cryosurgical eradication of the tumor was studied, using an experimental mammary tumor of the rat, MRMT-1. It was revealed that rejection rate of the challenged tumor increased gradually following cryosurgery and reached its peak at 10 weeks after cryosurgery. No such phenomenon was observed after surgical excision of the tumor. Decreased incidence of lymph node metastases and decreased tumor weights in “take” cases also suggested an increased immunological activity against the tumor at 10 weeks after cryosurgery.     

Immunological response of regional lymph nodes after tumor cryosurgery: experimental study in rats

Seven days after inoculation of metastasizing rat mammary tumor No. 1 into the thigh of 5-week-old female Sprague-Dawley rats, the tumor was treated cryosurgically by two-cycle freezing and by contact methods at -170 degrees C. Weights of the thymus and the spleen, histological findings of the lumbar lymph nodes, phytohemagglutinin (PHA)-induced blastogenesis of lymphocytes obtained from the lumbar lymph nodes and peripheral blood, and resistance rate to tumor rechallenge were examined 1, 3, 6, 10, and 17 week(s) after cryosurgery, with the following results: Thymus weight gradually decreased by 3 weeks after cryosurgery, while spleen weight increased by 1 week, recovering the preoperative level at 6 weeks. Paracortical hyperplasia of the lumbar lymph nodes markedly increased in 1 week and sinus histiocytosis increased after 3 weeks, both remaining at high values until 10 weeks, while germinal center hyperplasia showed a high value at 3 weeks and thereafter decreased gradually. PHA-induced blastogenesis of the lumbar lymph nodes significantly increased 1 week after cryosurgery and remained at its high value until 10 weeks. PHA-induced blastogenesis of peripheral lymphocytes showed the lowest value at 3 weeks and then significantly increased at 6 weeks. Resistance rate to rechallenge showed the lowest value at 3 weeks, reaching the highest level 10 weeks after cryosurgery. From the above results, it was suggested that anti-tumor immunity (resistance to tumor rechallenge) induced by cryosurgery was at the lowest level at 3 weeks after cryosurgery, and gradually increased starting at 6 weeks.(ABSTRACT TRUNCATED AT 250 WORDS)     

The sensitivity of experimental tumor systems to ultra low doses of doxorubicin

Biological effect of doxorubycin in standard (10(-3) mol/l) and ultra low doses (10(-5)-10(-20) mol/l) against some "signal" animal tumor systems--Lewis lung carcinoma, 755 adenocarcinoma, B-16 melanoma, Ehrlich carcinoma and L1210 leukemia was studied. The all models were very sensitive to the action of the drug in standard dose. Solid tumors' growth inhibition by 80-95% as well as increasing in life span of mice with L1210 leukemia by 86% in comparison with control and surviving of animals with Ehrlich carcinoma had been revealed. It had been shown that the drug in the area of ultra low doses occurred the following effects: inhibition of Lewis lung carcinoma growth by 80-95% compared to control after administration of the all tested ultra low doses; increasing of the life span of the animals with Ehrlich carcinoma and L1210 leukemia by 86-123% and 6-23%, correspondingly, upon the action of all tested ultra low doses; inhibition of B-16 melanoma growth by 50 and 70% after administration of the drug in doses 10(-20) mol/l and 10(-5) mol/l, correspondingly as well as deceleration of 755 carcinoma growth by 40% compared to control after action of the drug in the dose 10(-20) mol/l; stimulation of the B-16 melanoma growth by 20% relative to control after 10(-10) mol/l dose injection and enhancement of tumors sizes by 20-60% above control levels as a result of treatment of mice with 755 carcinoma by the drug in such ultra low doses as 10(-5) and 10(-15) mol/l. So, it was found that all tested tumor systems revealed certain sensitivity to the some ultra low doses of the drug. At the same time it was shown that doxorubycin in ultra low doses displayed alternative character of its biological effect, directivity of which varied according with the dose level and tumor strain.     

Current flow in skin frozen in experimental cryosurgery

Experiments were performed correlating tissue temperature and current flow during freezing canine skin. Current flow fell gradually to negligible levels as the tissue froze. Tissue temperature of ?31 °C correlated with current flow of 5 μA. Tissue temperature of ?55 °C correlated with zero current flow. The observations indicated that current flow was related to tissue temperature and that either or both could be used to predict tissue destruction in cryosurgery. However, tissue temperature measurement is the more accurate method of quantification.     

Critical temperature for skin necrosis in experimental cryosurgery

To investigate the minimal lethal freezing temperature required to produce skin necrosis in dogs, multiple skin sites were frozen with cryosurgical equipment. Tissue temperatures were recorded from thermocouple sites placed at diverse distances, usually 5 mm from the edge of the freezing probe. In single freezing cycles of about 3 min duration, tissue temperatures in the range of 0 to ?60 °C were produced. Punch biopsies of the skin at the thermocouple sites 3 days after freezing injury provided tissues for estimation of viability by histologic examination.The histologic findings permitted classification of the biopsy tissue into three groups, that is, viable, borderline, or necrotic. When classified as borderline, the division between the necrotic and viable tissue was evident on the histologic slide. The viable specimens were scattered through the 0 to ?35 °C range. All specimens frozen to ?10 °C or warmer were viable. In biopsies classified as borderline, the range of viability extended from ?11 ° to ?50 °C. The necrotic biopsies covered a range of ?14 ° to ?50 °C. Cell death was certain at temperatures colder than ?50 °C. The data showed cryosurgical freezing conditions produced a range of temperatures in which viability or death of tissue may occur and that the ranges of viability and necrosis overlapped to a great extent.The wide range of temperatures at which cells were viable shows the need to achieve tissue temperatures in the range of ?50 °C in the cryosurgical treatment of cancer.     

Tissue impedance and temperature measurements in relation to necrosis in experimental cryosurgery

Tissue temperature and impedance were measured in dog skin during freezing in situ. The previously frozen skin was removed by punch biopsies 3 days later to permit microscopic evaluation of the extent of necrosis. The histologic observations were related to the temperature and impedance measurements in an effort to determine the usefulness of the monitoring techniques in clinical cryosurgery. Tissue temperature and impedance have a definite relationship in tissue freezing, but the range of temperatures about any impedance values causes some concern. The tissue biopsies showed that an impedance value of at least 10 Mohms is not always associated with tissue death. In these experiments, there was the usual range of temperatures in relation to tissue death, but tissue temperatures of -30 degrees C and colder were always associated with complete necrosis. It is concluded that tissue temperatures are the more accurate and useful monitoring technique to supplement clinical judgment. However, impedance techniques may also be used to monitor therapy, especially if used primarily to monitor depth of therapy, and if controlled by clinical judgment wary of the inaccuracy of the technique.     

The experimental study for efficacy and safety of pancreatic cryosurgery

Objective: This study was designed to basic information concerning the efficacy and safety of cryosurgery for pancreatic cancer. Fifteen healthy pigs were used to perform biochemical analysis and histological assessment. Methods: Following anesthesia and laparotomy, an argon–helium cryoprobe was inserted into the pancreas. The introduction of argon gas induced a rapid decrease in temperature to ?160 °C (Group I, 5 pigs) or ?110 °C (Group II, 5 pigs), respectively, resulting in ice-ball formation of 15–20 mm diameter after 5 min. Following freezing, helium gas was circulated in the probe tip to increase the temperature to 10–20 °C over 3 min to thaw. The freeze/thaw cycle was then repeated. Group III (3 pigs) had a cryoprobe inserted, but without freezing, and Group IV (2 pigs) included untreated or normal control animals. Levels of serum amylase (AMY), IL-6 and C-RP were measured prior to freezing and for 7 days following the procedure. All pigs were euthanized 7 days post-treatment and pancreases were examined histologically. Results: Neither hyperaemia, edema or hemorrhage were observed in the un-frozen parts of the pancreas. Histological assessment revealed a significant level of necrosis in the central and lateral regions of the tissue frozen within the ice-ball. All cellular ultrastructure was destroyed and only observable as a few of remaining nuclei with broken crests and degranulated mitochondria and rough endoplasmic reticulum. There was a significant increase of serum AMY levels for a brief period in both “deep frozen” and the “shallow frozen” groups. However, the AMY also increased in two pigs in the “normal control” group and one pig from the “inserted cryoprobe without freeze” control group. All experimental pigs appeared healthy until the sacrifice time. Conclusion: Cryosurgery is a safe and effective ablative procedure for pancreatic tissue resulting in minimal complications.     

Effect of varying freezing and thawing rates in experimental cryosurgery

Six different freezing/thawing programs, which varied freezing rate, duration of freezing, and thawing rates, were used to investigate the effect of these factors on cell destruction in dog skin. The range of tissue temperatures produced was from -15 to -50 degrees C. The extent of destruction was evaluated by skin biopsies 3 days after cold injury. In single, short freezing/thawing cycles, the temperature reached in the tissue was the prime factor in cell death. Longer freezing time and slow thawing were also important lethal factors which increased destruction of cells. Cooling rate, whether slow or fast, made little difference in the outcome. The experiments suggested that present-day, commonly employed cryosurgical techniques, which feature fast cooling, slow thawing, and repetition of the freeze/thaw cycle, should be modified by the use of maintenance of the tissue in the frozen state for several minutes and slow thawing. Thawing should be complete before freezing is repeated. These modifications in technique will maximize tissue destruction, an important consideration in cancer cryosurgery.     

Numerical prediction of freezing fronts in cryosurgery: comparison with experimental results

Recent developments in scientific computing now allow to consider realistic applications of numerical modelling to medicine. In this work, a numerical method is presented for the simulation of phase change occurring in cryosurgery applications. The ultimate goal of these simulations is to accurately predict the freezing front position and the thermal history inside the ice ball which is essential to determine if cancerous cells have been completely destroyed. A semi-phase field formulation including blood flow considerations is employed for the simulations. Numerical results are enhanced by the introduction of an anisotropic remeshing strategy. The numerical procedure is validated by comparing the predictions of the model with experimental results.     

Effects of cryosurgery in experimental carcinoma on lectin binding and keratin distribution

Histochemical alterations of lectin binding and keratin distribution in experimental carcinomas of the hamster cheek pouch were obtained following cryotreatment. Cryotreated carcinoma cells showed a characteristic reduction in lectin binding and keratin staining shortly following cryosurgery. Tumor tissue, on the 2nd and 3rd days after cryotreatment, displayed destruction and necrosis with almost a complete loss of lectin binding and keratin staining. The remaining neoplastic cells located in the deeper layer showed positive reaction for both lectin binding and keratin, which is indicative of tumor recurrence. Histochemical staining of lectin binding and keratin proteins were useful markers in cryotreated tumor cells to identify either destruction and necrosis or vital activity of neoplastic growth.     

Response of completely mixed systems to pH shock

The response of aerobically growing heterogeneous microbial populations of sewage origin to step increases and decreases in pH were studied in both once-through and cell recycle systems. The pH range studied was 2.7 to 8.0. All studies were conducted at a dilution rate of 0.125 hr^?1, and all shocks were administered from a base or preshock pH level of 6.4 to 6.7. In each experiment, the preshock or initial “steady state” was assessed, the pH of the feed changed, and the resulting transient behavior of the system examined until attainment of the new or final “steady state” was approached. The major objectives of the work were to characterize the nature of the response with respect to biomass and effluent substrate concentrations, types of microbial populations present and chemical composition of the biomass, and to obtain guidelines as to allowable change in pH in waste streams. It was found in once-through systems that substrate removal efficiency recovered from pH levels as low as 3.0 after rather long periods of transient leakage of substrate. Cell recycle attenuated the severity of substrate leakage. In all cases of severe acid shock, the microbial population changed from predominantly bacterial-protozoan to one consisting predominantly of filamentous fungi. Changes in chemical composition of the sludge (protein and carbohydrate content) were consistent with the population changes. Based upon the results, it can be conservatively estimated that changes in pH of no more than one unit from the neutral preshock range can be tolerated without possible disruption of biochemical efficiency of substrate removal.     

Response of experimental epileptic focus to focal ionizing radiation

An experimental model of cobalt-induced epilepsy was performed on 15 cats. 3 months later, they were irradiated by means of a gamma source with a total target dose of 10 Gy. One month after irradiation all EEG recordings returned to normal, while a control group maintained the epileptiform traces. Histological study of the irradiated foci showed neuronal reafferentation. The action mechanism of ionizing radiation over epileptogenic foci in relation to this reafferentation and neuronal plasticity mechanisms is discussed.     

Two freezing cycles ensure interface sterilization by cryosurgery during bone tumor resection.

Cryosurgery utilizing an argon-based system allows bone-tumor interface sterilization, while avoiding the risks of conventional cryosurgery. This study was conducted in order to evaluate the number of freezing cycles required for interface sterilization in cases of aggressive human bone tumors. Sixteen tumors were included (six chondrosarcomas, eight metastatic carcinomas, and two giant cell tumors). All occurred within long bones. In all cases a standardized marginal resection was performed. Following thorough curettage, we sampled five different locations within the tumor interface by a cylindrical hollow trephine. The interface viability was assessed using the XTT method. Quantitative histological evaluation was based on the percentage of live cells divided by total lacunae number in five random medium-power fields. One freezing cycle (5 min, -40 degrees C) reduced tumor viability to approximately 5% of prefreezing. However, there were still live specimens. Two or three freezing cycles led to complete interface sterilization. The difference between a single freezing cycle and two freezing cycles was significant (ANOVA, F = 130, P < 0.01). The difference between two freezing cycles and three freezing cycles was not significant (ANOVA, F = 0.14, P < 0.6). The results of the XTT method for the assessment of interface viability correlated well with histological evaluation of the percentage of viable cells (r = 0.89), as well as with cell culture results of frozen vs. prefreezing tumor samples. In conclusion, two freezing cycles are adequate to achieve tumor-bone interface sterilization in aggressive human bone tumors.